Applications of isothermal titration calorimetry - the research and technical developments from 2011 to 2015

Abstract

Isothermal titration calorimetry is a widely used biophysical technique for studying the formation or dissociation of molecular complexes. Over the last 5 years, much work has been published on the interpretation of isothermal titration calorimetry (ITC) data for single binding and multiple binding sites. As over 80% of ITC papers are on macromolecules of biological origin, this interpretation is challenging. Some researchers have attempted to link the thermodynamics constants to events at the molecular level. This review highlights work carried out using binding sites characterized using x‐ray crystallography techniques that allow speculation about individual bond formation and the displacement of individual water molecules during ligand binding and link these events to the thermodynamic constants for binding. The review also considers research conducted with synthetic binding partners where specific binding events like anion‐π and π‐π interactions were studied. The revival of assays that enable both thermodynamic and kinetic information to be collected from ITC data is highlighted. Lastly, published criticism of ITC research from a physical chemistry perspective is appraised and practical advice provided for researchers unfamiliar with thermodynamics and its interpretation. Copyright © 2016 John Wiley & Sons, Ltd.

Biophysical characterization of antibodies with isothermal titration calorimetry

“…The lowest K D value (tightest binding affinity) that can be directly measured by ITC is in the single-digit nanomolar range [5][6][7][8][9][10][11]. ITC data for tighter affinities (single digit nanomolar and lower) cannot be precisely determined from a single ITC titration experiment, since the binding will be saturated within one or two injections, and there are not enough data points to calculate the K D from the binding isotherm.…”

A Thermodynamic Model of Auto‐regulated Protein Assembly by a Supramolecular Scaffold

“…Isothermal titration calorimetry (ITC) is the method of choice for thermodynamic characterization of molecular interactions ,. Importantly for auto‐regulated assembly, both ligand binding and assembly processes yield heat changes that can be detected by ITC.…”

“…Isothermal titration calorimetry (ITC) is the method of choice for thermodynamic characterization of molecular interactions. [25,26] Importantly for auto-regulated assembly, both ligand binding and assembly processes yield heat changes that can be detected by ITC. For standard ITC analysis a single set of independent binding sites model is fit to the experimental data with the affinity and enthalpy parameters extracted simultaneously by non-linear least squares methods.…”

Critical assessment of relevant methods in the field of biosensors with direct optical detection based on fibers and waveguides using plasmonic, resonance, and interference effects

“…Since that year, the number of publications has increased, and publications cover especially the field of protein chemistry. Research and technical development from 2011 to 2015 has been reviewed, providing information on methodological advances and interpretation of single and multiple binding sites [70]. Besides thermodynamic information on binding constants, e.g., enzymes, of substrate reactions and inhibitory constants, kinetic data is also of interest.…”